Introduction―Tissue factor (TF), the main blood coagulation cascade activator, has been proposed to be present, among circulating cells, also in platelets. The presence of both protein and mRNA in platelets suggests that the source of TF could be through several mechanisms: microparticle-transfer, megakaryocyte-transfer or endogenous synthesis. Despite these evidences, the issue of platelet-associated TF and of its presence also in megakaryocytes is still matter of debate because some other investigator fail to detect it. Aim―To prove that megakaryocytes as well as platelets contain both TF mRNA and protein using a megakaryoblastic cell line cultured in vitro, in order to avoid the cross-talk with the others circulating cells. Methods―MEG01 were cultured with valproic acid (VPA) to induce differentiation and platelet-like particles (PLPs) production. Megakaryocytes and PLPs were characterized for the presence of specific population markers (CD41/CD61) and for TF expression by flow-cytometry and immunocytochemistry. TF mRNA was assessed by RT-PCR both in megakaryocytes and PLPs. Results―VPA-induced megakarypoiesis of MEG01 resulted in an increased percentage of adherent cells (from 2,6±1,5 to 70±2,3) which showed the typical feature of megakaryocytes such as polyploidy, formation of proplatelets and elevated expression of the megakaryocytic marker CD41 and CD61 (90% and 95%). PLPs were also CD61+ (65%) and CD41+ (30%). TF protein was expressed by both megakaryocytes and PLPs(47,4%±1,4 and 14,5±5,7). Interestingly, stimulation of PLPs with A23187 resulted in an increase in the number of TF-positive cells(25%±3). RT-PCR confirmed the presence of TF as well as of GpIIIa, GpIb and COX1 mRNA both in megakaryocytes and in PLPs. Conclusions―The use of MEG01 allowed us to prove the presence of TF mRNA and protein both in megakaryocytes and PLPs supporting the hypothesis that the megakaryocyte-transfer mechanism may account for the presence of TF in platelets.
|Titolo:||Platelet-associated Tissue factor expression : shedding light on a controversial issue|
|Parole Chiave:||tissue factor ; platelets ; megakaryocyte ; valproic acid|
|Settore Scientifico Disciplinare:||Settore BIO/14 - Farmacologia|
|Data di pubblicazione:||set-2012|
|Appare nelle tipologie:||01 - Articolo su periodico|